exoUmutant on variousL. japonicusecotypes
revealed that both host and environmental factors
were linked to the requirement for EPS. These
results reveal a complex function forM. lotiEPS
in determinate nodule formation and suggest that
EPS plays a signaling role at the stages of both IT
initiation and bacterial release (Kelly et al. 2013 ).
Genes for LPS synthesis may also have a role
in nodulation. At least some LPS mutants of
M. lotiform nodules impaired in nitrogenfixa-
tion (Turska-Szewczuk et al. 2009 ), while others
form nitrogen-fixing nodules but are impaired in
competitive ability (D’Antuono et al. 2005 ). The
purified lipid A moiety of M. loti strain
MAFF303099 LPS is able to induce nitrous
oxide production inL. japonicusroots, suggest-
ing that the plant is able to recognize rhizobial
LPS (Hashimoto et al. 2012 ; Murakami et al.
2011 ). However, a precise role for rhizobial LPS
in the nodulation of Lotus species is yet to
be defined.M. lotimutants defective in cyclic
ß-glucan synthesis are also defective in nodule
invasion, with a possible role of the cyclic
ß-glucan being in osmotic protection of the rhi-
zobia (D’Antuono et al. 2005 ; Kawaharada et al.
2007 , 2010 ).
5.4 Common Properties ofM. loti
Symbiosis Islands
To date, sequences of symbiosis islands from
M. lotiMAFF303099, R7A, and NZP2037 have
been published. The three islands share highly
collinear regions with multiple deletions and
insertions (Fig.5.2; see also Sullivan et al. 2002 ).
The shared DNA regions contain genes for
nodulation, nitrogenfixation, conjugal transfer of
the island, and metabolic enzymes. The sizes of
islands are approximately 502, 533, and 611 kb
in R7A, NZP2037, and MAFF303099, respec-
tively (Kaneko et al. 2000 ; Kasai-Maita et al.
2013 ; Sullivan et al. 2002 ). The number of genes
they harbor is proportional to their size, about 1
gene/kbp: 414, 504, and 583 genes. As shown for
R7A, the island in MAFF303099 is also inserted
as an entity in the phe-tRNA gene, with a 17-bp
duplication of the 3′-portion demarcating the
other end of the island. The island of NZP2037
seems to have similarly inserted in a phe-tRNA
gene; however, it has been split into two parts of
528 and 5 kb, through a recombination event
likely mediated by a transposon. The collinear
regions of three islands share 165 genes among a
total of 1,078 non-redundant genes. These con-
served genes, or core genes, on the symbiosis
islands substantially cover the genes for nodula-
tion, nitrogenfixation, and energy conversion in
nodules as listed in Table5.1. The numbers of
genes shared by two of the three strains are 33,
25, and 35, respectively, by MAFF303099–R7A,
MAFF303099–NZP2037, and R7A–NZP2037
pairs. The rest of the genes, 820 in total, are
strain specific and mostly genes of unknown
function and transposon-related genes such as
transposases, integrases, and resolvases (Kasai-
Maita et al. 2013 ). Nevertheless, some of the
strain-specific genes include genes related to Nod
factor synthesis and protein secretion systems
that are involved in the interaction with host
legumes.5.4.1 Genes Involved in Nod Factor
Synthesis and ExportAmong the core genes on the symbiosis islands,
there are 16nod,noe, andnolgenes involved in
the synthesis, modification, and excretion of the
lipochito-oligosaccharides (LCOs) or Nod fac-
tors that trigger the host developmental program
to form nodules (Kaneko et al. 2000 ; Kasai-
Maita et al. 2013 ; Sullivan et al. 2002 ). These
include two copies of the regulatory genenodD,
the commonnodgenesnodA, B,andC(with
nodBin a separate operon fromnodAC, unlike
other rhizobia) as well as other genes involved in
chemical modifications of the core LCO. The
onset of signal exchange generally takes place
whenflavonoids or isoflavones secreted by the
host legume are bound by the rhizobial NodD
protein that then activates transcription of other46 K. Saeki and C.W. Ronson